Resistance to collagen-induced arthritis in a nonhuman primate species maps to the major histocompatibility complex class I region.

Type II collagen-induced arthritis (CIA) is an experimentally inducible autoimmune disorder that is, just like several forms of human arthritis, influenced by a genetic background. Immunization of young rhesus monkeys (Macaca mulatta) with type II collagen (CII) induced CIA in about 70% of the animals. One major histocompatibility complex (MHC) class I allele was present only in young animals resistant to CIA and absent in arthritic animals. This strong association suggests that the MHC class I allele itself, or a closely linked gene, determines resistance to CIA. The mechanism controlling the resistance to CIA becomes less efficient in aged animals since older rhesus monkeys, which were positive for the resistance marker, developed a mild form of arthritis. At the cellular level it is demonstrated that resistance to CIA is reflected by a low responsiveness of T cells to CII. This association between a specified MHC class I allele and resistance to an autoimmune disease points at the importance of the MHC class I region in the regulation of the immune response to an autoantigen.

Antiarthritic activity of the newly developed neutrophil oxidative burst antagonist apocynin.

The plant-phenol 4-hydroxy-3-methoxyacetophenone (trivial name apocynin) is a strong inhibitor of neutrophil superoxide anion (O2-) release in vitro. In vitro the inhibitory effect of apocynin is restricted to cells with the capacity to release peroxidase and reactive oxygen species (ROS). Peroxidase deficient cells are insensitive to apocynin. In the present study the antiinflammatory activity of apocynin was tested in collagen-induced arthritis in rats. Collagen-immunized rats were treated with different doses of apocynin in the drinking water starting at the onset of joint-swelling and terminating 14 days later, at the time when joint swelling in the control group was maximal. Apocynin-treated animals had a normal plasma level of collagen-specific antibodies, but showed a significant reduction of the joint swelling. Also the plasma IL-6 level in apocynin-treated animals was substantially lower than in control animals. No flare-up of joint swelling after termination of the treatment was observed in the apocynin-treated groups.

Attenuation of malaria infection, paralysis and lesions in the central nervous system by low protein diets in rats.

Young Wistar rats developed a fulminant infection when inoculated with the rodent malaria parasite Plasmodium berghei. Rats that died during the infection exhibited a progressive paralysis of the extremities, a rapidly decreasing body temperature and minute haemorrhages in the brain. Increasing the level of protein in the diet from 4 to 8 and 16% was accompanied by an increase in morbidity and mortality from 15 to 40 and 90% respectively on day 6 of the infection. Increasing the level of dietary protein also increased the reticulocyte count of the peripheral blood in infected and non-infected rats. The attenuation of the cerebral syndrome in rats fed a diet low in protein may be related to changes in erythropoiesis or to changes in immune reactivity.

The newly developed neutrophil oxidative burst antagonist apocynin inhibits joint-swelling in rat collagen arthritis.

Methylated catechols like apocynin selectively inhibits neutrophil superoxide production without significant side-effects. When rats were treated orally with low doses of apocynin the severity of collagen-induced arthritis was significantly reduced, pointing at a role of oxyradicals in the induction of this disease.

The anti-arthritic and immunosuppressive effects of cyclosporin A on collagen-induced arthritis in the rhesus monkey.

The influence of cyclosporin A (CsA) on type II collagen-induced arthritis (CIA) in the rhesus monkey has been investigated. CsA was administered subcutaneously in a dose of 25 mg/kg per day during 9-18 days and additionally 12.5 mg/kg per day for 7 days. At this dosing regime no significant alterations of haematologic parameters were found, indicating that the toxicity of CsA was negligible. Administration of CsA after onset of arthritis had no beneficial effect, but when given between immunization and manifestation of clinical symptoms, CIA could be prevented completely. Moreover, these monkeys became resistant to the disease, because no arthritic activity could be observed upon a booster immunization with type II collagen (CII). The suppression of disease by CsA is reflected in reduced antibody levels to CII.

Resistance to collagen-induced arthritis in rats and rhesus monkeys after immunization with attenuated type II collagen.

Immunization of susceptible rodent or primate species with type II collagen (b-CII) from bovine origin induces type II collagen-induced arthritis (CIA). The disease is characterized as a systemic polyarthritis associated with humoral and cellular autoimmunity to CII and shares similarity with human arthritic diseases. The objective of this study was to develop a procedure for induction of resistance to CIA in animals, which possess a certain major histocompatibility complex phenotype that makes them prone to develop CIA (susceptible). It is shown that by immunization with an attenuated form of CII, in which arthritogenic epitopes have been destroyed by heat denaturation, disease resistance is induced in a susceptible inbred rat strain (RT-1u) and in an outbred population of susceptible rhesus monkeys (lacking the Mamu-A26 allele). In both species the disease resistance is connected with modulation of anti-CII autoantibodies of the IgM isotype. This protocol may provide a basis for effective and safe methods to induce protection to autoimmune arthritis in those subjects that are genetically prone to develop such a disease.

The mitogen-induced generation of interferon-gamma producing cells in cultures of rhesus monkey peripheral blood mononuclear cells is age-dependent.

An enzyme-linked immunoabsorbent spot (ELISPOT) assay has been developed for measuring the frequency of interferon-gamma (IFN-gamma) producing cells in rhesus monkeys. Aged monkeys revealed, upon mitogenic stimulation, a significantly higher percentage of IFN-gamma secreting peripheral blood mononuclear cells (PBMC) compared to young animals. No correlation was found between the frequency of IFN-gamma producing PBMC and the mitogen-driven proliferation, indicating that in rhesus monkeys no direct correlation exists between these two activation parameters.

Acquired resistance to type II collagen-induced arthritis in rhesus monkeys is reflected by a T cell low-responsiveness to the antigen.

Ten out of 14 rhesus monkeys developed arthritis after a single immunization with bovine type II collagen (B-CII). In contrast to primary resistant monkeys, arthritic animals showed a B-CII specific T cell proliferation during the induction phase of the disease. All surviving animals showed a full remission of the disease. Two monkeys acquired resistance to collagen-induced arthritis (CIA) after one period of disease, but in three animals a booster immunization with B-CII induced a slight flare-up. It is demonstrated that B-CII immunized rhesus monkeys have the capacity to restore resistance to CIA. The development of resistance to CIA is reflected by a decreased T cell responsiveness to B-CII. It is shown that the lack of IL-2 plays a role in B-CII-induced T cell low-responsiveness. A potential role of CD8+ T cells in the down regulation of the T cell response to B-CII is discussed.

Induction of type II collagen-specific antibody production in blood lymphocyte cultures of rhesus monkeys (Macaca mulatta) with collagen-induced arthritis using the immobilized native antigen.

Peripheral blood mononuclear cells (PBMC) from Rhesus monkeys previously immunized with bovine type II collagen to induce arthritis were cultured with the same antigen. Because the native protein is poorly soluble in culture medium a heating step is often used. The antigen in this form induced PBMC proliferation, but epitopes for the induction of antibody production and arthritis were lost. To keep the native protein intact it was coated on affigel beads. With the immobilized antigen specific antibody production could be induced.

Identification of antibody epitopes in the CB-11 peptide of bovine type II collagen recognized by sera from arthritis-susceptible and -resistant rhesus monkeys.

Sera from eight rhesus monkeys that had been immunized with native bovine type II collagen were tested for antibodies to cyanogen bromide peptides (CB peptides) of type II collagen by Western blotting. The monkeys produced IgG antibodies to a number of different CB peptides, with five out of eight animals producing antibodies to the CB-11 peptide (four arthritic, one non-arthritic). Antibody epitopes on the CB-11 peptide of bovine type II collagen recognized by these sera were investigated by epitope mapping. Peptides (8-mers overlapping by seven amino acids) representing the CB-11 region were synthesised and the sera screened for binding to these peptides to determine areas of high IgG antibody binding to this region of type II collagen. The profiles obtained were not identical, though there were some epitopes that were commonly recognized. Antibodies to one epitope, also present in human type II collagen, were found only in the sera of two animals with the severest arthritis. The technique of epitope mapping has successfully identified a number of epitopes within the CB-11 peptide of type II collagen recognized by antibodies from bovine type II collagen-immunized monkeys. Studies on the relevance of responses to the identified epitopes can now be undertaken.

Collagen-induced arthritis in an outbred group of rhesus monkeys comprising responder and nonresponder animals. Relationship between the course of arthritis and collagen-specific immunity.

It is speculated that the autoimmune response to type II collagen (CII) is a driving force in the pathogenesis of human rheumatoid arthritis (RA). In this report, we describe the relationship between the induction of collagen arthritis and the CII-specific humoral, as well as cellular, immune response in rhesus monkeys. Ten of 14 monkeys immunized with bovine type II collagen (B-CII) developed polyarthritis. Susceptible animals showed a T cell response to B-CII; resistant animals did not. After the primary immunization, the humoral response to B-CII, as well as to rhesus monkey type II collagen, was dominated by antibodies of the IgM isotype in the susceptible animals and by antibodies of the IgG isotype in the resistant animals. Because of the close phylogenic relationship between the rhesus monkey and humans, these data contribute valuable information about the role of CII-specific immunity in the pathogenesis of human RA.

Induction of an anti-vaccine response by T cell vaccination in non-human primates and humans.

Experimental and spontaneous autoimmune disease in animals can effectively be prevented and treated by application of pathogenic autoreactive T cells in an attenuated form. This approach has become known as T cell vaccination. T cell vaccination exploits specifically the ability of the immune system to regulate its autoreactive T cells by mechanisms of network control. The success of T cell vaccination in a variety of rodent animal models has raised hopes for its use as an effective and specific therapy in human autoimmune disease. The aim of this study was to induce an anti-T cell response by T cell vaccination in humans and primates as a pre-clinical study into the feasibility and toxicity of T cell vaccination. Using bulk cultures of T cells from the peripheral blood or an inflamed joint, it was possible to induce a T cell response specific for the injected vaccine and its activation state both in rhesus monkeys and in two patients with active rheumatoid arthritis. In one of the patients there was already a spontaneous T cell response against a mitogen driven T cell line from the peripheral blood, but not against a control T cell line specific for tetanus toxoid, suggesting that regulatory T cell networks are operative in patients with autoimmune disease. Significant clinical effects or side-effects were not observed. The results suggest that T cell vaccination in humans is feasible and non-toxic. It is likely to influence an already ongoing regulatory process. Conditions for making T cell vaccination an effective therapy need still to be worked out by further studies both in primates and in less complex human immune processes.

Experimental immune mediated arthritis in rhesus monkeys. A model for human rheumatoid arthritis?

The induction of experimental arthritis in rhesus monkeys was studied by intradermal immunization of bovine type II collagen and antigens derived from Mycobacterium tuberculosis, Streptococcus pyogenes, and Eubacterium aerofaciens. The tested bacterial antigens proved to be not arthrogenic. Bovine type II collagen induced clinical arthritis in 50% of the rhesus monkeys. Type II collagen induced arthritis in rhesus monkeys proved to be a potential model to study clinical, serological, histological, genetic, and immunologic features associated with human RA.